DESCRIPTION: The long term goal is to develop methods for clinical application of gene therapy to human neurodegenerative diseases. This proposal focuses on ex vivo and in vivo gene therapy to provide the potent neurotrophic factor, GDNF, to dopamine (DA) neurons in the Parkinsonian (PD) brain model. Herpes simplex amplicon (HSV), adenoviral (Ad) and retroviral (RV) vectors will be made for GDNF , a frameshift mutant GDNF as a control, and the cellular marker gene, nuclear localized LacZnl. Vectors will be made containing each of 3 cellular promoters, the chicken B-actin constitutive, the neuronal promoter, NSE, or the astrocyte promoter, GFAP, or viral promoters. Promoter specificity, strength and duration of expression in the context of HSV, Ad, and RV vectors will be compared in primary cultures of neurons and glia, and in vivo in rat brain. Vectors will be used for in vivo and ex vivo gene therapy in well characterized rat models of PD to study effects of GDNF gene therapy on damaged DA neurons in the adult brain and fetal DA neurons grafted into adult brain. A battery of behavioral tests of DA-dependent striatal functions will be used to compare efficacy of the different approaches. The effects of gene therapy with GDNF will also be compared to those of injecting recombinant GDNF into the brain. In addition, HSV vectors harboring the neuroprotective proto-oncogene, bcl-2, under control of the tyrosine hydroxylase promoter will be made and used in vivo to determine whether expression of bcl-2 in DA neurons an adult brain rescues them from damage induced by neurotoxin MPP+. Methods in molecular biology, immunocytochemistry, neuromorphometry, primary neuronal culture, Elisa assay, bioassay, rodent microsurgery and behavioral testing will be applied. These studies are directly relevant to clinical intervention in humans suffering from PD and to clinical trials utilizing fetal grafts. More generally, they promise to increase our knowledge on the potential of gene therapy to treat neurodegenerative diseases and methods for obtaining long term gene transgene expression in CNS from viral vectors.